Asthma is a prevalent chronic inflammatory disease characterized by constriction of the airways. Beta-adrenergic receptors (Beta2AR) on bronchial smooth muscle relax this constriction and are the targets of beta-agonists used for treatment bronchospasm. However, there is a high degree of inter-individual variation in beta2AR function due to factors that include receptor regulation by the asthmatic milieu, beta-agonist treatment, and polymorphisms of the receptor. Thus the most commonly utilized class of drugs for the treatment of asthma is likely suboptimal because of deficiencies in our understanding of target signal regulation. The long-term objective is to understand how beta2AR signaling is regulated by genetic factors, agonist structure and modulation of post- receptor transduction elements. In Specific Aim 1, the combinations of polymorphisms of the beta2AR gene found in the human population, arranged as haplotypes, will be studied within the context of expression and agonist regulation. In Specific Aim 2, the molecular basis of pleiotropic responses to agonist will be delineated. Here, we will delineate specific properties of agonists with respect to coupling to Gs, Gi and MAP kinase, sequestration, phosphorylation. and down- regulation using a multiply recombinant approach. In Specific Aim 3, the role of Gi expression in beta2AR signaling in the airway will be determined. Two transgenic mice will be created over-expressing Gi or a Gi inhibitor in airway smooth muscle. Since an increase in Gi has been reported to be an important component of beta2AR and bronchial hyper- reactivity in asthma, we will be able to directly assess this potential mechanism in isolation of other processes. The approach includes biochemical, ex vivo, and in vivo studies of beta2AR function in cells, trachea and intact mice so as to merge the in vitro results with relevant physiological function. In Specific Aim 4, the rate limiting step of beta2AR signal transduction will be explored by generating two transgenic mice, over-expressing adenylyl cyclase type VI or a peptide inhibitor, in airway smooth muscle. Again, a comprehensive approach linking signaling events and physiology will be utilized. These studies will further our understanding of beta2AR regulation and provide new insights into asthma pathophysiology and treatment.